The structural details of Golgi membranes and their associated vesicles are still undefined. Both the continuity between adjacent Golgi sacs and the relationship between the trans-golgi cisternae and the network of vesicles that emanates from it (the TGN) are unresolved. We are using selective stains for this region of the cell, including photo-converted fluorescent dyes, antibodies, and enzymatic markers, to render the trans-golgi region electron dense and suitable for detailed fine structural study. We have prepared 10 tomograms from chemically fixed cells that have been stained with labeled ceremide to reveal the trans Golgi cisternae and the TGN. These tomograms reveal clathrin coated vesicles and others with a novel coat morphology forming in adjacent regions of the trans Golgi network (Ladinsky et al., 1994). We are now studying cells fixed by rapid freezing and freeze-substitution to get information about membrane arrangements in cells preserved by optimum methods. Preliminary work with these specimens confirms our identification of different morphologies on vesicles budding from the TGN and suggests unexpected relationships among the successive cisternae in the Golgi stack. Issues of membrane continuity between stacks and between Golgi elements and endosomes are now being examined in this well preserved material.